Modification of the GeneScan 2500 fluorescent dye standard for accurate product sizing.
AuthorsBartlett JM, Crilly A, White A, Madhok R
JournalMol Biotechnol
PubMed ID10934531
'This article describes a procedure for modification of the commercially prepared GeneScan 2500 size standard for allelotyping with large DNA fragments such as variable number tandem repeats (VNTRs). Here a procedure was used to adapt commercially available size standards for the sizing of the interleukin-6 (IL6) 3''VNTR, which has allele ... More
A new class of homogeneous nucleic acid probes based on specific displacement hybridization.
AuthorsLi Q, Luan G, Guo Q, Liang J
JournalNucleic Acids Res
PubMed ID11788731
'We have developed a new class of probes for homogeneous nucleic acid detection based on the proposed displacement hybridization. Our probes consist of two complementary oligodeoxyribonucleotides of different length labeled with a fluorophore and a quencher in close proximity in the duplex. The probes on their own are quenched, but ... More
A novel method for real time quantitative RT-PCR.
AuthorsGibson UE, Heid CA, Williams PM
JournalGenome Res
PubMed ID8908519
'A novel approach to quantitative reverse transcriptase polymerase chain reaction (QC RT-PCR) using real time detection and the 5'' nuclease assay has been developed. Cystic fibrosis transmembrane transductance regulator (CFTR) target mRNA is reverse transcribed, amplified, detected, and quantitated in real time. A fluorogenic probe was designed to detect the ... More
High-throughput analysis of telomerase by capillary electrophoresis.
AuthorsAtha DH, Miller K, Sanow AD, Xu J, Hess JL, Wu OC, Wang W, Srivastava S, Highsmith WE
JournalElectrophoresis
PubMed ID12652580
'The enzyme telomerase is expressed in (85-90)% of all human cancers, but not in normal, non-stem cell somatic tissues. Clinical assays for telomerase in easily obtained body fluids would have great utility as noninvasive, cost-effective methods for the early detection of cancer. The most commonly used method for the detection ... More
Convenient genotyping of six myostatin mutations causing double-muscling in cattle using a multiplex oligonucleotide ligation assay.
AuthorsKarim L, Coppieters W, Grobet L, Valentini A, Georges M
JournalAnim Genet
PubMed ID11167526
We herein describe a procedure that allows for simultaneous genotyping of six loss-of-function mutations in the bovine myostatin gene associated with the double-muscling phenotype. The proposed method relies on a multiplex oligonucleotide ligation assay and detection of the fluorescently labelled products using automatic sequencers. ... More
Typing of the short tandem repeat D8S347 locus with different fluorescence markers.
AuthorsPöltl R, Luckenbach C, Fimmers R, Ritter H
JournalElectrophoresis
PubMed ID9504824
The short tandem repeat (STR) locus D8S347 was analyzed by capillary electrophoresis. Sequencing data and a population study of 203 individuals from a southwestern German population are presented. We detected 12 different alleles, 340-388 bp in length, and found 40 different genotypes. The heterozygosity index was 85.7%. Futhermore, we investigated ... More
Efficiencies of fluorescence resonance energy transfer and contact-mediated quenching in oligonucleotide probes.
AuthorsMarras SA, Kramer FR, Tyagi S
JournalNucleic Acids Res
PubMed ID12409481
An important consideration in the design of oligonucleotide probes for homogeneous hybridization assays is the efficiency of energy transfer between the fluorophore and quencher used to label the probes. We have determined the efficiency of energy transfer for a large number of combinations of commonly used fluorophores and quenchers. We ... More
Chromosome-specific panels of tri- and tetranucleotide microsatellite markers for multiplex fluorescent detection and automated genotyping: evaluation of their utility in pathology and forensics.
A set of 391 microsatellite markers (Weber set 6), 85% of which consist of tri- and tetranucleotide repeat markers, were used to design chromosome-specific panels that allowed for a high degree of multiplexing with respect to the fragment size range and fluorophore (FAM, HEX, TET). This marker set has an ... More
Single nucleotide polymorphism genotyping using short, fluorescently labeled locked nucleic acid (LNA) probes and fluorescence polarization detection.
AuthorsSimeonov A, Nikiforov TT
JournalNucleic Acids Res
PubMed ID12202779
Locked nucleic acids (LNAs) are synthetic nucleic acid analogs that bind to complementary target molecules (DNA, RNA or LNA) with very high affinity. At the same time, this binding affinity is decreased substantially when the hybrids thus formed contain even a single mismatched base pair. We have exploited these properties ... More
Single nucleotide polymorphisms (SNPs) are becoming widely recognized as the new currency for gene mapping as increasing numbers are discovered. Here we outline a method for their rapid analysis based on an allele-specific polymerase chain reaction (PCR) which employs a competitive approach, whereby both allele-specific primers are present in the ... More
Influence of fluorophor dye labels on the migration behavior of polymerase chain reaction--amplified short tandem repeats during denaturing capillary electrophoresis.
AuthorsHahn M, Wilhelm J, Pingoud A
JournalElectrophoresis
PubMed ID11545394
The determination of the length of polymerase chain reaction (PCR)-amplified short tandem repeats (STRs) by denaturing capillary electrophoresis (CE) is a standard procedure for purposes of genotyping. We show that dye-specific mobility anomalies exist for 5'-fluorophor-labelled single-stranded DNA (ssDNA) fragments in CE using the performance-optimized polymer 4 (POP4) buffer sieving ... More
Rapid identification of up to three Candida species in a single reaction tube by a 5' exonuclease assay using fluorescent DNA probes.
AuthorsShin JH, Nolte FS, Holloway BP, Morrison CJ
JournalJ Clin Microbiol
PubMed ID9854084
We used fungus-specific PCR primers and species-specific DNA probes to detect up to three Candida species in a single reaction tube by exploiting the 5' to 3' exonuclease activity of Taq DNA polymerase. Probes to the internal transcribed spacer region of the rRNA gene were labeled at the 5' end ... More
Novel algorithm identifies species in a polymycobacterial sample by fluorescence capillary electrophoresis-based single-strand conformation polymorphism analysis.
AuthorsIwamoto T, Sonobe T, Hayashi K
JournalJ Clin Microbiol
PubMed ID12454176
An algorithm to directly identify multiple mycobacterial species in a sample by using fluorescence capillary electrophoresis (FCE)-based single-strand conformation polymorphism (SSCP) analysis was developed. Part of the 16S-23S ribosomal DNA internal transcribed spacer (ITS) region in 37 reference strains and 73 clinical isolates representing 19 mycobacterial species and Mycobacterium tuberculosis ... More